The reelin signaling pathway has been studied for over 50 years as a critical regulator of cell positioning and migration in the developing central nervous system. My laboratory has recently identified a novel role for reelin signaling in regulating mammary gland morphogenesis. Using immunohistochemical approaches and reporter gene expression, we have shown that components of the reelin signaling pathway are expressed in the developing and mature mammary gland and that alterations in reelin signaling cause improper ductal branching and disrupt epithelial organization in vivo, resulting in abnormal development of the mammary ductal network. We have also shown that isolated mammary epithelial cells, which line the ductal lumen, slow their migration in the presence of reelin protein. These observations lead to the hypothesis that reelin signaling may be a critical regulator of cell migration in the mammary gland. This proposal will address how Dab1, an intracellular adaptor protein that is activated in response to reelin signaling, functions in regulating the behavior of mammary epithelial cells. Several isoforms of Dab1 have been identified in embryonic brain and retina; these isoforms may allow a differential response to the presence of reelin and a nuanced response to reelin signaling. My laboratory has demonstrated the expression of several distinct Dab1 isoforms in the mammary gland and this proposal will determine the biological functions of these isoforms. First, we will determine when and where the different isoforms are expressed, with the expectation that isoforms expressed in proliferating and migrating mammary epithelial cells will be different from isoforms expressed in quiescent cells. Second, we will determine if changing isoform expression alters the behavior of mammary epithelial cells both in vitro and in vivo. Third, we will determine which elements of the reelin signaling pathway are activated by the different Dab1 isoforms in mammary epithelial cells. There are several alternative outcomes to reelin signaling in neuronal cells and we expect that isoform expression may dictate which elements of the pathways are activated in mammary epithelial cells. Together, these experiments will demonstrate that differential expression of Dab1 isoforms is important for the regulation of cellular behaviors in the mammary gland. Understanding the mechanisms that regulate mammary epithelial cell migration may ultimately lead to therapeutic approaches to inhibit the metastatic migration of breast cancer cells.